Peroxisomes are metabolic compartments that are essential for human and plant development. Although understanding of how matrix proteins are imported into peroxisomes is increasing, mechanisms for turning over damaged or obsolete peroxisomal proteins remain largely obscure. The proposed studies will address peroxisome-associated protein degradation in Arabidopsis thaliana; the unique peroxisomal functions and facile genetics of this system will allow dissection of peroxisomal processes in an intact organism. Peroxins, encoded by PEX genes, are necessary for peroxisome biogenesis. Most peroxins act in matrix protein import; this work will determine whether a subset of these peroxins have additional roles in matrix protein turnover. Preliminary data suggest that a subset of peroxins work together both in receptor recycling and in a novel type of retrotranslocation - the removal for degradation of peroxisomal matrix proteins that are damaged or no longer needed. Three specific aims are proposed to identify components and molecular requirements of peroxisome-associated protein degradation. Aim 1 will characterize matrix protein localization and stability in mutants defective in peroxins and proteasome subunits. Aim 2 will identify cis-acting molecular signals necessary for degradation of two peroxisomal matrix proteins that undergo regulated destruction during a specific stage of seedling development. Aim 3 will recover mutants defective in known and undiscovered components of the peroxisome-associated degradation machinery. The successful completion of these aims will begin to address an unsolved mystery of peroxisome biology: How do peroxisomes dispose of damaged or obsolete proteins? Peroxisomal defects underlie a group of inherited syndromes known as peroxisome biogenesis disorders, which are generally fatal in infancy and are characterized by severe mental retardation, neuronal migration defects, craniofacial abnormalities, and other symptoms. The proposed experiments will exploit unique aspects of plant peroxisomes while taking advantage of knowledge from fungal and mammalian systems to provide insights that are likely to apply throughout eukaryotes. Adding an evolutionarily distinct model to the study of peroxisome biology will allow the development of model and hypotheses to expand and refine our understanding of these essential organelles. PUBLIC HEALTH RELEVANCE: Using Arabidopsis to Uncover New Roles for Peroxins Peroxisomes are subcellular compartments housing critical metabolic reactions and are essential for normal human development. Peroxisomal defects underlie a group of inherited syndromes known as peroxisome biogenesis disorders, which are generally fatal early in infancy. The proposed experiments will elucidate how cells dispose of proteins within peroxisomes when they are damaged or no longer needed.